T. cruzi and other important human pathogens establish chronic infections after mucosal invasion, making both mucosal and systemic immune studies relevant for the development of protective vaccines. In our current RO1, we hypothesized that Th1 and Th2 cells would be differentially important for systemic and mucosal immunity, respectively, and mutually inhibitory. Contrary to our original hypotheses, we have shown that T. cruzi specific Th1 responses provide both optimal mucosal and systemic protection, strongly supporting the feasibility of vaccines designed to induce both optimal mucosal and systemic immunity concurrently. Further challenges remain, however. First, another important route of T. cruzi infection is through breaks in the skin. Safe and effective molecular vaccines capable of inducing mucosal, cutaneous and systemic T. cruzi protection must be developed. Second, although Th1 responses are more protective than Th2 responses, Th1 responses may not induce optimal secretory IgA also mechanistically important for mucosal protection. Third, the mechanisms involved in trafficking of vaccine-induced memory T cells to mucosal, cutaneous and systemic sites of T. cruzi infection are not known. We plan to further study T. cruzi as a model for investigations of the molecular and cellular relationships between mucosal, cutaneous and systemic immunity. We will study 3 major hypotheses: 1) Cruzipain and/or trans-sialidase specific immunity can protect against mucosal, cutaneous and systemic T. cruzi infection without inducing immunopathology, 2) Th1 biased vaccines induce serum IgG and secretory IgA responses protective against T. cruzi mucosal and cutaneous infection, and 3) Specific chemokine receptors and other trafficking molecules expressed on memory T cells are important for T. cruzi mucosal, cutaneous and systemic protection. Recombinant protein, DNA and live vaccines will be studied alone and in prime/boosting combinations. We will directly assess the protective effects of serum IgG, secretory IgA and B cells against mucosal and cutaneous challenges. Chemokine and/or integrin receptor expression will be studied by flow cytometry and RT-Real-Time PCR, and the in vivo effects of defective chemokine and integrin binding and/or signaling will be evaluated.